Studies will be continued with Pronase endopeptidases to determine what groups are regulating the pH dependence for activity in the alkaline range. These will include quantitative titration studies and also an examination of those groups which are modified by acetic anhydride in the absence of glycerol which result in loss of activity. The studies on Pronase and bovine trypsins will be continued to examine if homoarginine residues are selectively cleaved by the mammalian enzyme. The studies on quanidinated proteins will be extended so that exchange-in studies with tritium will be complemented by exchange-out studies. In addition the stabilities of the guanidinated enzymes as compared to their native precursors will be followed in increasing concentrations of urea or guanidine; conformational changes will be followed by susceptibility to proteolysis by the very stable Pronase endopeptidases, UV absorption, and measurements of circular dichroism. The studies on human trypsin and human trypsin inhibitor will be extended to see how human trypsin inhibitor interacts with Pronase trypsin and what the effects of pH, metal ions, and ionic strength are upon that interaction. Studies will be continued with the primary structures of Pronase carboxypeptidase and aminopeptidase. In addition efforts will be continued to resolve and characterize the higher molecular weight components of Pronase; to date these proteins have not been successfully resolved by the usual purification techniques.